This proposal requests funds to purchase a new super-resolution optical microscope, the OMX Blaze V4 from Applied Precision, to enhance the NIH-funded research projects of Major Users at Vanderbilt University. Recent developments in imaging technology have made it possible to resolve objects in biological samples with a size below the diffraction limit of the conventional light microscope (~200 nm). From the perspective of a cell biologist, this is a critical breakthrough; proteins, protein complexes, vesicles and othr small organelles are all in the sub-200 nm size regime. Thus, super-resolution optical microcopy now makes it possible to directly observe these fine sub-cellular features and precisely localize even single molecules in the context of intact fixed and live cells. The OMX Blaze V4 system from Applied Precision enables users to take advantage of two super-resolution imaging modes: structured illumination microscopy (SIM) for fast simultaneous multi- color super-resolution imaging of live cells (lateral resolution = 80-100 nm), and localization microscopy (LM) which provides the highest lateral resolution possible for the imaging of fixed samples (lateral resolution = 20- 50 nm). Major Users (80% usage) are members of the newly formed Epithelial Biology Center (EBC, co- directed by Bob Coffey, M.D. and Jim Goldenring, M.D. Ph.D.); these investigators will apply super-resolution imaging to break new ground in their NIH-funded research projects, which span a wide range of problems in cell and developmental biology, epithelial biology, cancer biology and microbiology. Projects include studies of cytoskeletal dynamics (Kaverina, Ohi, Tyska), polarized membrane trafficking (Coffey, Crowe, Goldenring, Kaverina), mitotic spindle assembly and function (Ohi), assembly of cell surface features such as microvilli and cilia (Goldenring, Tyska), and host-pathogen interactions (Crowe, Tyska). Acquisition of this system will also benefit a group of Minor Users (20% usage) investigating complementary problems at the subcellular level (Janetopoulos, Kenworthy, Miller, Prince, Webb, and Weaver). Together, these investigations hold significance for understanding disease and illness related to birth defects, cancer, bacterial/viral infections, and many other human health problems. The proposed OMX microscope will be installed in the EBC and managed by the Vanderbilt Cell Imaging Shared Resource (CISR, managed by Sam Wells, Ph.D.). With the help of substantial support from the Vanderbilt Office of Research, the CISR will provide the infrastructure for introducing researchers across Vanderbilt (~275 laboratories) to this exciting new technology, training users in the proper operation of this instrument, and applying an established business model to support the long-term maintenance of this system. In summary, acquisition of the OMX will provide unprecedented cellular imaging capabilities that currently do not exist on the Vanderbilt campus. This instrument will enable investigators to image cells with a spatial resolution approaching molecular dimensions, pushing research of biological problems in directions that are simply not possible with the microscopes currently available to Vanderbilt users.